Common Carp Populations Research Articles

Development of a novel droplet digital PCR assay for the sensitive detection of carp edema virus

Carp edema virus (CEV) is a poxvirus associated with outbreaks of clinical disease that may lead to high mortality in both koi and common carp populations, potentially causing severe economic losses for carp aquaculture and the global koi trade. Therefore, the establishment of a sensitive and specific assay for detection of CEV is an essential requirement for the prevention of further spread of this emerging viral disease. In this study, a novel droplet-digital PCR (ddPCR) assay, targeting the p4a gene of CEV was developed. The assay exhibits good linearity, repeatability and reproducibility. Linearity was maintained at extremely low concentrations of CEV nucleic acid templates. The detection limit of ddPCR was 2.2 ± 0.26 copies of CEV DNA per reaction (n = 8), which represents a greater sensitivity than the quantitative PCR (qPCR) assay. Specificity analysis showed that the ddPCR assay for CEV had no cross-reactivity with other important aquatic pathogens. In clinical diagnosis of 151 koi and common-carp samples, 58 and 45 tested positively by ddPCR and qPCR assays, respectively. The overall agreement between the two assays was 91.39% (138/151). Linear regression analysis demonstrated a significant correlation between ddPCR and qPCR results with an R2 value of 0.9627. Our findings indicate that the ddPCR assay could provide a robust diagnostic tool for the sensitive detection of CEV, even in samples with a low viral load.

Genetic Investigation of Aral Wild Common Carp Populations (Cyprinus carpio) Using ddRAD Sequencing

Common carp (Cyprinus carpio) is a widespread freshwater fish species of the Cyprinidae family, one of the largest and most diverse fish families. The natural habitats of C. carpio extend from Western Europe to South-East Asia. Common carp has remained an economically important fish species in aquaculture for many centuries and its production nowadays exceeds 4 million tons worldwide and continues to grow. The taxonomy of C. carpio is complicated, since this species is usually distinguished in two, three, and even four distinct subspecies. In the present study, we used ddRAD-sequencing to genotype 30 specimens from five wild common carp populations from the Ponto-Caspian, Balkhash-Ile, and Aral Sea geographical regions. It is demonstrated that they differ at the population level according to F-statistics analysis. At the same time, the subspecies status of C. carpio aralensis has not yet been confirmed. We found several loci that can be used as a discriminant for Aral and Ponto-Caspian wild common carp populations. It is suggested that Aral carp (C. carpio aralensis), which inhabits Balkhash-Ile and Aral Sea basins, is related to Ponto-Caspian or European carp (C. carpio carpio). Moreover, Aral carp might be the ancestor for European carp subspecies. Our results can be used to develop population-specific, high-density SNP marker panels, allowing the trade control of common carp production in the Eurasian Economic Union.

Genetic contribution of domestic European common carp (Cyprinus carpio carpio) and Amur carp (Cyprinus carpio haematopterus) to the wild Vietnamese carp population as revealed by ddRAD sequencing

At least three subspecies of the common carp, namely, Cyprinus carpio carpio, C. carpio haematopterus, and C. carpio viridiviolaceus, have been described to date. These subspecies natively inhabit Ponto-Caspian and Far Eastern regions, as well as northern Vietnam, but they have also been introduced into aquaculture in almost every region of the world where the environmental conditions are suitable. Moreover, common carp is one of the most widely cultured freshwater fish species in the world. In northern Vietnam, common carp fisheries and aquacultural production are of great economic importance. Investigation of the subspecies distribution among aquacultured and wild common carps is important for understanding the impact of human economic activity, such as aquaculture production, on the environment. In the present study, double-digest restriction site-associated DNA sequencing (ddRAD-seq) was used to estimate the genetic impact of aquaculture carps on the wild common carp populations of northern Vietnam. Our work reveals that a part of wild carp populations has a significant contribution to farmed fish ancestry. For some populations, this contribution exceeds 25% of the population ancestry, but the ancestry value is distributed across most specimens of the populations, whereas pure aquaculture specimens are quite uncommon for wild habitats.

Some population parameters of the common carp (Cyprinus carpio, 1758) in Lake Paniai, Papua

Common carp (Cyprinus carpio) is known as introduced fish on some lakes in Indonesia and included in herbivore fish. The common carp population in Lake is the dominant number, so that it’s become at of fishing target. Estimation of some parameters of common carp population in Lake Paniai was aimed to evaluate the growth parameters, mortality, the fishing rate and recruitment pattern. Collecting and measuring the fish sample was done in the period from February to October 2016 by using the fishing gear of gill nets with mesh sizes of 1.00 - 4.50 inches. The results showed that common carp population in Lake Paniai was dominated by individual lengths between 15-25 cm with a frequency of 55.78%, the growth pattern of male fish was allometric (-) and females were isometric. Asymptotic length (L∞) = 61.43 cm and the growth coefficient (K) = 0.32 per year. The rate of natural mortality (M) = 0.65 per year, the fishing mortality rate (F) = 0.52 per year, the total mortality rate (Z) = 1.17 per year and the exploitation rate (E) = 0.44. The exploitation rate of common carp fish was still below the optimum value, thus an attempt to catch this fish, could be still improved. Common carp populations caught by mostly fishermen have had a chance to spawning thus common carp fish population in the lake was estimated to still be preserved and could be utilized in a sustainably. Peak recruitment occurred only once in a year, namely in June.

Simulated Use of YY Male Stocking and Suppression for Eradicating Common Carp Populations

AbstractThe introduction and subsequent spread of Common CarpCyprinus carpiothroughout the world represent one of the most destructive and pervasive forces impacting aquatic ecosystems. Herein we use computer simulations to predict if stocking YY male carp in conjunction with manual netting or piscicide use could eradicate invasive Common Carp populations. Two forms of YY males were evaluated, including sperm‐producing (MYY) and egg‐producing (FYY) fish. A stochastic individual‐based population model was used to evaluate the effects of simulated management actions on both the abundance and extirpation probabilities of high‐ and low‐density populations. Results predicted that some YY male stocking scenarios alone could eradicate carp populations. For example, when stocking FYYcarp into low‐density populations, predicted extirpation probabilities exceeded 0.87 when stocking at least 50 carp/ha was maintained for at least 10 years. Additionally, a one‐time use of rotenone (to reduce the virtual population by 90%) plus stocking at least 50 FYYcarp/ha for at least 5 years into low‐density populations resulted in predicted extirpation probabilities exceeding 0.88. Favorable extirpation probabilities were also obtained by combining YY male stocking and either selective (i.e., no culling of stocked YY males) or non‐selective annual suppression. For example, FYYstocking with only 10% sustained, non‐selective suppression yielded extinction probabilities exceeding 95% in 15 years, while increasing suppression levels to 50% resulted in extirpation probabilities over 95% within 5 years. Without exception, MYYstocking functioned far less efficiently in simulations than FYYstocking, but MYYcarp did work in a small subset of low‐density scenarios and may also have prophylactic utility where prior eradication has occurred and where there is a high risk of re‐invasion. We conclude that stocking of YY male Common Carp is worthy of field testing, but additional simulations are needed to explore other questions not addressed in this study.

Interrelationships among bluegill demographics, predator and competitor abundances and impoundment characteristics

Bluegills Lepomis macrochirus R. provide popular sport fisheries but the quality of these fisheries has declined across much of their range. Factors that may influence bluegill demographics were examined in 18 Midwestern (USA) impoundments which contained largemouth bass Micropterus salmoides L., the main apex predator, as well as varying abundances of gizzard shad Dorosoma cepedianum L. and common carp Cyprinus carpio L., which can negatively influence the quality of bluegill populations. Regression models revealed that the relative abundance (catch-per-unit-effort [CPUE], number per hour of electrofishing) of bluegill was positively associated with largemouth bass CPUE and total phosphorous concentration. The total lengths (TL) and ages of the largest and oldest bluegills were negatively associated with common carp CPUE and total annual mortality of bluegills, and positively associated with the TL and age of bluegills when 50% were sexually mature and largemouth bass CPUE. Total annual mortality negatively influenced the TL and age at sexual maturity. Bluegills matured at larger sizes and older ages in impoundments with clearer water. This study revealed the strong influence of mortality on size and age structure and the onset of sexual maturity of bluegill, and the negative influence of common carp on these populations. Management strategies to improve bluegill populations should involve reducing bluegill mortality and common carp populations.

Molecular characterization of carp edema virus disease: An emerging threat to koi Cyprinus carpio in China

Carp edema virus disease (CEVD) has resulted in great economic losses in koi (Cyprinus carpio koi) and common carp (Cyprinus carpio carpio) populations in the world. In this study, the diseased koi were diagnosed as CEV infection based on 5′ untranslated region (5′UTR) and 4a protein genes by the conventional PCR, nested PCR and quantitative PCR (qPCR) analyses. Phylogenetic tree analysis showed that the TJ201708 strain was classified into the genogroup IIa. Furthermore, qPCR of 5′UTR gene revealed that the lowest detection limit was 4.0 fg/μL. The pathogenicity of CEV for koi was demonstrated in the infection experiments. Histopathological examination revealed the petechial hemorrhages of liver and spleen, vacuolization of lamina propria of intestine and swelling and necrosis of respiratory epithelial cells of gills. To our knowledge, this is the first report the qPCR of 5′UTR gene in the detection of carp edema virus.

Analysis of the spawning population of common carp in the lower reaches of the Ural River within the borders of the West Kazakhstan Region

The article presents the results of a study of the hydrological regime of the Ural river in the territory of the West Kazakhstan region, the average annual flow of the river, as well as the average decadal water levels during the spring flood. According to the presented results, there is a noticeable decrease in annual runoff, which directly has a negative impact on the local ichthyofauna of the reservoir. The decrease in the water level in the Ural river affects the efficiency of natural reproduction of phytophilic fish. In 2019, due to a decrease in the water content of the Ural river, carp fish, namely common carp, did not have the opportunity to spawn in floodplains and flood meadows of the river. When catching producers for the purpose of artificial reproduction, the spawning herd of common carp was represented by individuals aged from 5 to 12 years.

A new strain group of common carp: The genetic differences and admixture events between Cyprinus carpio breeds.

Common carp (Cyprinus carpio) has an outstanding economic importance in freshwater aquaculture due to its high adaptive capacity to both food and environment. In fact, it is the third most farmed fish species worldwide according to the Food and Agriculture Organization. More than four million tons of common carp are produced annually in aquaculture, and more than a hundred thousand tons are caught from the wild. Historically, the common carp was also the first fish species to be domesticated in ancient China, and now, there is a huge variety of domestic carp strains worldwide. In the present study, we used double digestion restriction site‐associated DNA sequencing to genotype several European common carp strains and showed that they are divided into two distinct groups. One of them includes central European common carp strains as well as Ponto–Caspian wild common carp populations, whereas the other group contains several common carp strains that originated in the Soviet Union, mostly as cold‐resistant strains. We believe that breeding with wild Amur carp and subsequent selection of the hybrids for resistance to adverse environmental conditions was the attribute of the second group. We assessed the contribution of wild Amur carp inheritance to the common carp strains and discovered discriminating genes, which differed in allele frequencies between groups. Taken together, our results improve our current understanding of the genetic variability of common carp, namely the structure of natural and artificial carp populations, and the contribution of wild carp traits to domestic strains.

Patterns of Geographical and Potential Adaptive Divergence in the Genome of the Common Carp (Cyprinus carpio).

The common carp, Cyprinus carpio, is a cyprinid fish species cultured in Europe and Asia. It accounts for >70% of freshwater aquaculture production worldwide. We conducted a population genomics analysis on C. carpio using high-throughput SNP genotyping of 2,198 individuals from 14 populations worldwide to determine the genetic architecture of common carp populations and the genetic bases for environmental adaptation. Structure analyses including phylogeny and principal component analysis were also conducted, showing distinct geographical patterns in European and Asian populations. The linkage disequilibrium block average lengths of the 14 populations ranged from 3.94 kb to 36.67 kb. Genes within selective sweep regions were identified by genome scanning among the different populations, including gdf6a, bmpr1b, and opsin5. Gene Ontology and KEGG enrichment analyses revealed potential trait-related loci and genes associated with body shape, scaling patterns, and skin color. This population genomics analysis may provide valuable clues for future genome-assisted breeding of C. carpio.

Population Characteristics and the Potential Suppression of Common Carp in Lake Spokane, Washington

Abstract Common Carp Cyprinus carpio is a nonnative species that often has deleterious effects on aquatic systems. As such, there is interest in suppressing nonnative Common Carp populations in areas where humans have introduced them. The objectives of this study were to 1) provide insight on efficient techniques for capturing Common Carp, 2) describe their population demographics and dynamics, 3) evaluate whether temperature and water elevation were related to growth and recruitment, and 4) develop an age-structured population model for evaluating different management scenarios of Common Carp removal in Lake Spokane, Washington. Catch rates of Common Carp varied among sampling gears with slightly higher catch rates in monofilament (mean ± SD; 15.5 ± 9.8 fish/net night) vs. multifilament (12.7 ± 7.3 fish/net night) gill nets. Catch rates of Common Carp with nighttime electrofishing (0.3 ± 0.4 fish/min) were higher than daytime electrofishing (0.1 ± 0.2 fish/min). Common Carp in Lake Spokane exhibited variable recruitment, rapid growth, large-length structure, high longevity (i.e., age 18 y), and low total annual mortality (17.0%). Air temperature was positively associated with annual growth increments (R2 ≤ 0.25). Neither air temperature nor water elevation was highly correlated (R2 ≤ 0.20) to recruitment of Common Carp. A Beverton–Holt yield-per-recruit model suggested that yield declined with increasing exploitation. Recruitment overfishing would occur at exploitation rates of 20–40% for all targeted minimum length categories (i.e., 150, 300, 450 mm) except 600 mm. Results from this study provide important information on the ecology of Common Carp that can be used to guide management efforts (e.g., suppression) in western systems.

Genetic analysis for six wild and selection populations of common carp ( Cyprinus carpio ) using microsatellites

Genetic analysis for six wild and selection populations of common carp ( Cyprinus carpio ) using microsatellites

Effects of 17á-ethinylestradiol-based contraceptives on ovarian tissue using histopathological examination and FSH, LH, CYP1A, and CYP3A expressions using qRT-PCR in the ovaries and brain tissues of common carp (Cyprinus carpio L.)

In this study, the main aim was to identify the effects of exposure to 17á-ethinylestradiol-based contraceptives on the ovaries and brain tissues of common carp. For this purpose, histopathological changes were evaluated in ovaries after 21 days of contraceptive exposure. In addition, the expression levels of the CYP1A, CYP3A, FSH, and LH genes were measured in contraceptive-exposed brain tissue and ovaries respectively, using qRT-PCR (p less than 0.05, p less than 0.01, and p less than 0.001, respectively). No histopathological lesions, including inflammation, degeneration, or necrosis in the evaluated ovarian tissue of common carp were found after both high and low levels of exposure to contraceptives. Therefore, it is concluded that these contraceptives were highly dangerous to the reproduction system of common carp; the accumulation of contraceptive waste in the water supply could adversely affect and threat the common carp population. This may indirectly also affect other animals.

Analysis of complete genome and pathogenicity studies of the spring viremia of carp virus isolated from common carp (Cyprinus carpio carpio) and largemouth bass (Micropterus salmoides): An indication of SVC disease threat in Korea

A batch of wild common carp and largemouth bass died in Andong, Gyeongsangbuk-do province, South Korea, in 2016. Moribund fish showed typical signs of spring viremia of carp (SVC) disease, which causes acute hemorrhage in the skin and ascites. Thus far, SVC disease has been detected in several regions of the world but never in South Korea. Suspecting the infectious agent to be the SCV virus (SVCV), the moribund fish were sampled and screened. The isolated virus developed a cytopathic effect in EPC cells. Both viral isolates from the common carp (ADC-SVC2016-1) and largemouth bass (ADC-SVC2016-3) were identical in terms of their genome sequence, which were 11,034 bp nucleotides in length. Genome comparison exhibited greater sequence similarity with the Asian SVCV sequences available at NCBI. Phylogenetic analysis revealed that the Korean SVCV isolates were clustered within the Asian clade. More specifically, evolutionary analysis by using the P gene sequences showed that the Korean isolates were sub-cladded within the Iai genogroup but diverged from Chinese strains of SH150514 and SH160901. The Korean isolates shared more than 98% sequence similarity with the two Chinese SVCV isolates, suggesting that the spread of SVCV originated from China. The isolated virus had cytopathic effects on EPC cells. Virus transmission studies showed that the virus exhibited the highest virulence at 15 °C, which was also dependent on the method used, with the injection method being better than the immersion and cohabitation methods. This is the first study to document that Korean SVCV isolates may be epizootic in wild common carp and other susceptible animal populations in South Korea.

Evaluation of a new select-breeding carp variety with high fillet quality

The morphological characters, fillet texture, and nutrient contents of the F3 generation of Barbless carp (Cyprinus pellegrini,BC) female × Heilongjiang carp (Cyprinus carpio,HLC, geographical population of common carp) male hybrid, which were named as High-quality carp F3 (HQC F3), were studied. The main morphological characteristics of HQC F3 were between BC and HLC, but closer to the BC. The fillet amino acids and crude protein showed significant differences between HQC F3 and BC. But, only fillet crude fat showed significant differences between HQC F3 and HLC. HQC F3's fillet had significantly different with BC's on the contents of myristic acid, linolenic acid, and stearic acid, and with HLC's on the contents of linoleic acid, linolenic acid, EPA, palm oleic acid, and 20 carbon-dilute acid. HQC F3 broodstocks were selected with high intramuscular fat and effective population size. HQC F4 were bred and showed improved intramuscular fat content compared to F3. The intramuscular fat of 2-year-old HLC F4 was significantly (p < 0.05) correlated with body weight/standard length. Intramuscular fat is a stable trait that can be continuously passed on to the next generation. This study will lay a foundation for further carp selective breeding study to improve fillet quality.

Source–sink dynamics explain the distribution and persistence of an invasive population of common carp across a model Midwestern watershed

Source–sink theory is an ecological framework that describes how site and habitat-specific demographic rates and patch connectivity can explain population structure and persistence across heterogeneous landscapes. Although commonly used in conservation planning, source–sink theory has rarely been applied to the management of invasive species. This study tested whether the common carp, one of the world’s most invasive species, exhibits source–sink dynamics in a representative watershed in the Upper Mississippi River Basin comprised of a dozen interconnected ponds and lakes. To test for source–sink population structure, we used standard fish sampling techniques, tagging, and genetic assignment methods to describe habitat-specific recruitment rates and dispersal. Five years of sampling revealed that while adult carp were found across the entire watershed, reproductive success (the presence of young carp) was restricted to shallow ponds. Additionally, nearly a third of the carp tagged in a representative pond dispersed into the connected deeper lakes, suggesting that ponds in this system serve as sources and lakes as sinks. This possibility was confirmed by microsatellite analysis of carp tissue samples (n = 1041) which revealed the presence of two distinct strains of carp cohabitating in the lakes, whose natal origins could be traced back to one of two pond systems, with many adult carp attempting to migrate back into these natal ponds to spawn. We conclude that the distribution and persistence of invasive carp in complex interconnected systems may often be driven by source–sink dynamics and that their populations could be controlled by suppressing reproduction in source habitats or by disrupting dispersal pathways, instead of culling individuals from sink habitats.

Assessing the efficacy of corn-based bait containing antimycin-a to control common carp populations using laboratory and pond experiments

Strategic use of oral toxicants could allow for practical and sustainable control schemes for the invasive common carp (Cyprinus carpio, or ‘carp’) if a toxicant selectively targeted carp and not native species. In this study, we incorporated antimycin-a (ANT-A), a known fish toxicant, into a corn-based bait and conducted a series of experiments to determine its toxicity, leaching rate, and species-specificity. Our results showed that ANT-A was lethal to carp at doses ≥ 4 mg/kg and that the amount of ANT-A that leached out of the bait in 72 h was not lethal to carp or bluegill (Lepomis macrochirus). Species-specificity trials were conducted in 227 L tanks, in which carp were stocked with three native species representing families that occur sympatrically with carp in our study region: the fathead minnow (Pimephales promelas), yellow perch (Perca flavescens) and bluegill. These trials showed high mortality of carp (46%) and fathead minnows (76%) but no significant mortality of perch or bluegill. Finally, a pond study, which used the same species composition except for fathead minnows, resulted in 37% morality among adult carp and no mortality among perch or bluegill. Our results suggest that corn-based bait that contains ANT-A could be used to selectively control carp in ecosystems dominated by percids or centrarchids, such as lakes across the Great Plains ecoregion of North America, where carp are especially problematic.

Impacts of an invasive virus (CyHV-3) on established invasive populations of common carp (Cyprinus carpio) in North America

The effects of invasive pathogens on wild fish and fish communities generally are not well documented. We compiled information on the impacts of mass mortality events due to Cyprinid Herpesvirus-3 (CyHV-3), otherwise known as Koi Herpesvirus, on wild North American populations of the invasive cyprinid, Cyprinus carpio (common carp), based on our personal experiences, discussions with North American fish ecologists and virologists, a detailed survey of technical and popular publications and a web search. We found evidence of 17 mass die-offs of carp due to CyHV-3 in North America since 2004, for 7 of which we were able to obtain information about carp before and after the events. For 6 of the events, effects of the die-offs on carp population indices appeared to be slight. Carp size-frequency distributions before and after the well-documented 2007/08 event in Ontario were also not conspicuously different. The exceptional event was at Blue Springs Lake, Missouri, in 2012, at which we estimate 65% of the carp present died as a result of CyHV-3 infections and carp abundance continues to decline. Why Blue Springs Lake differs from other events in North America is not clear. Overall, carp die-offs due to CyHV-3 in North America (1) confirm laboratory studies that only common carp are affected, (2) are of brief duration (3–6 weeks), (3) are not repeated in subsequent years and (4) cause much lower mortality (with the exception of Blue Springs Lake) than previously reported for carp in aquaculture facilities or in the laboratory. In terms of both wild carp and their effects on aquatic communities, the short and long-term effects of most die-offs appear to be slight. These features could have implications for the effectiveness of the proposed use of CyHV-3 to reduce feral carp populations in Australia.

Historical demography of common carp estimated from individuals collected from various parts of the world using the pairwise sequentially markovian coalescent approach.

The inference of historical demography of a species is helpful for understanding species' differentiation and its population dynamics. However, such inference has been previously difficult due to the lack of proper analytical methods and availability of genetic data. A recently developed method called Pairwise Sequentially Markovian Coalescent (PSMC) offers the capability for estimation of the trajectories of historical populations over considerable time periods using genomic sequences. In this study, we applied this approach to infer the historical demography of the common carp using samples collected from Europe, Asia and the Americas. Comparison between Asian and European common carp populations showed that the last glacial period starting 100ka BP likely caused a significant decline in population size of the wild common carp in Europe, while it did not have much of an impact on its counterparts in Asia. This was probably caused by differences in glacial activities in East Asia and Europe, and suggesting a separation of the European and Asian clades before the last glacial maximum. The North American clade which is an invasive population shared a similar demographic history as those from Europe, consistent with the idea that the North American common carp probably had European ancestral origins. Our analysis represents the first reconstruction of the historical population demography of the common carp, which is important to elucidate the separation of European and Asian common carp clades during the Quaternary glaciation, as well as the dispersal of common carp across the world.

Histopathology and Detection of Cyprinid Herpesvirus Infection in Two Koi Farms in Tianjin City (China)

Koi herpesvirus (KHV) is the etiologic agent of koi herpesvirus disease (KHVD) causing mass mortalities in common carp (Cyprinus carpio carpio), koi (Cyprinus carpio koi), and ghost carp (Cyprinus carpio goi) populations. In this study, we report new outbreaks of the koi herpesvirus (KHV) disease in a koi carp farm in Tianjin city, northern China. From June to September 2017, severe mortalities of koi juvenile and broodstock occurred in ornamental fish, and infected koi carp which showed signs of lethargy and loss of appetite, and clinical signs including sunken eyes, gill necrosis, and excessive secretion of mucus. Histopathologically, the gills showed hyperplasia and degeneration of epithelial cells and fusion of the lamellae, and foci necrosis was found in gills, liver, kidney, and spleen. In addition, KHV and Carp edema virus (CEV) were detected in the diseased koi fish by diagnostic PCR tests for the viruses. The results showed that the nucleotide sequences of thymidine kinase (TK), DNA polymerase (SphI-5), and four open reading frame (ORF 25, ORF 56, ORF 72, ORF 81) genes confirmed the highest identity with other KHV strains such as KHV-I, KHV-U and KHV-J, but the CEV was negative. The phylogenetic tree showed KHV-TJ1708 and KHV-TJ1709 clustered with the Asian strain KHV-J (AP008984). Results of the present study confirmed the prevalence of Carp herpes disease (KHVD) in China koi populations by means of clinical examination, histopathology and molecular technique. These data will provide a reference for diagnosis, quarantine, and surveillance of KHV in China.